2.7. mtROS
The MitoSOX™ Red mitochondrial superoxide indicator (M36008; Invitrogen)
was used to detect mitochondrial ROS (mtROS) accumulation. The cells
were incubated with 1.0 mL of 5 μM MitoSOX™ reagent working solution for
10 min at 37°C in dark. Thereafter, the cells were washed gently with
warm buffer thrice, and fluorescence was observed and photographed under
a fluorescence microscope (DMI4000B; Leica, Wetzlar, Germany).